Name of Applicant (Last, First, Middle): GNATENKO, Dmitri V. ABSTRACT This SBIR Phase I application is submitted in response to Small Business Topics of Special Interest (TOSI) initiative HLS17-03. The overall goal of the proposal is to develop and commercialize an assay for detection of RNA biomarkers in circulating blood platelets, developed as a novel diagnostic approach with potential applicability to wide-ranging disorders including cancer, cardiovascular or autoimmune disorders. Platelets are anucleate blood cells that have a crucial role in the maintenance of hemostasis, thrombosis and wound healing. They contain a broad variety of miRNAs and mRNAs. In addition to their canonical functions, platelets mediate intercellular RNA transfer, and are known to have altered genetic signatures that allow for disease tracking and/or early diagnostics. We have successfully adapted and compared several RNA profiling platforms to transcript profiling of platelets and developed a class prediction model that discriminates clonal from non-clonal phenotypes of thrombocytosis with 93.6% accuracy. We will now extend these advances to combine fluorescent in situ hybridization and flow cytometry for multiplexed mRNA and miRNA biomarkers in individual platelets. The high sensitivity and specificity of this technology bypasses the need for technically rigorous RNA isolation, and allows for routine genetic biomarker detection and quantification using standard flow cytometric analyses. In Phase I we propose to develop a sensitive assay for detection and quantification of miRNA and mRNA transcripts in platelets at a single-cell level and to compare this technology to quantitative RT-PCR. Phase II will be focused on refinement, analytical and clinical validation of the technology for diagnostics and disease tracking human disorders encompassing genetically- altered biomarker subsets. Since platelets are easily accessible, this technology represents a significant step forward towards minimally invasive detection and quantification of RNA biomarkers in circulating blood.